1. Reference to Copending Patent Application
Reference is made to co-pending application Ser. No. 08/188,946 filed Jan. 27, 1994, entitled "IN VIVO DOSIMETER FOR PHOTODYNAMIC THERAPY" which is a continuation of U.S. patent application Ser. No. 07/786,036 filed Oct. 31, 1991, naming one of the present inventors (Daniel R. Doiron) as the inventor.
2. Prior Art
In photodynamic therapy and many other medical diagnostic and therapeutics applications it is important to measure either the relative or absolute concentration of fluorescent exogenous chromophore in tissues. Such a measurement can be used to diagnose a disease, such as cancer, viral inflection, vascular plaque, etc., or it can be used for determining the therapeutic dosage of a photosensitive drug such as in photodynamic therapy, chemotherapy, or radiation therapy using a radiation sensitizer. Doing such an in vivo fluorescence measurement in a repeatable and/or quantitative manner can be difficult due to a number of variables affecting the magnitude of the fluorescence signal obtained. These variables include:
(i) Tissue optical properties; PA1 (ii) Excitation wavelength extinction coefficient of the exogenous chromophore and intensity of excitation light; PA1 (iii) Fluorescence properties of the exogenous chromophore including: PA1 (iv) Relative geometry of the excitation and detection systems; PA1 (v) Tissue endogenous chromophore fluorescence interference (also know as autofluorescence or AF); and PA1 (vi) Gain characteristics of the detection system.
(a) Fluorescence quantum yield; PA2 (b) Binding site; PA2 (c) molecular environment conditions such as temperature and pH;
Many methods have been developed to measure the fluorescence intensity of various chromophores for analytical and diagnostic applications. To overcome the problems noted above, most methods for measuring the fluorescence intensity employ a very rigidly defined geometry requiring control of all parameters effecting the measured signal. For example, typical analytical fluorometers use a standard 90 degree illumination geometry for excitation and detection while using a fixed detection cell path length. The excitation intensity is monitored and ratioed to the measured fluorescence signal. This prior art ratioing method only corrects for variation of the excitation source while not providing any correction for the other parameters. A 90 degree system is not easily adapted for use with partially opaque or turbid materials.
For in vivo measurement of fluorescence a variety of methodologies may be used. One of the most common employs fiber optics and an Optical Multichannel Analyzer (OMA). Such a system generally uses one fiber to deliver the excitation light to the target tissue (tissue under investigation) and one or more fibers to collect and deliver the fluorescence light to the OMA. The OMA uses a diffractive or dispersion grating to spread the light out over a multichannel charge coupled device (CCD). The signal measured in a specific channel of the CCD can then be related to a specific wavelength, (or a narrow band of wavelengths). Such a system provides the general fluorescence emission spectrum of the tissue, but the intensity and shape of this curve will depend on many of the same parameters outlined above. Relating such a spectrum to the level of a specific chromophore requires the normalization of the spectrum along with some detailed spectral analysis to determine the portion of the fluorescence signal which is signal is due to the chromophore of interest. This is particularly difficult if the chromophore of interest is exogenous and there are similar endogenous chromophores present. The flurorescence spectra intensity and curve shape will vary significantly from tissue to tissue samples having either the same or different histology.
Recently a great deal of work has been published and patented on the use on in 9 vivo fluorescence spectroscopy of endogenous chromophores to determine the histological or pathological state of a particular tissue without taking a tissue biopsy. Such methods are based on the difference between the shape and/or intensity of the fluorescence emissions spectrum between normal tissue and diseased tissue. While such methods have been used for diagnosing and/or locating diseased tissue, they have not been applied to in vivo kinetic studies of photosensitive drugs in a particular tissue. It is of particular interest to determine the pharmacokinetics of a particular exogenous chromophore in vivo.